Growth factor delivery system for the healing of wounds and the prevention of inflammation and disease

ABSTRACT

The present invention features hydrogel drug delivery systems and methods of producing and using such systems for the treatment of wounds. The systems are based on a hydrogel into which a low concentration of growth factor, e.g., epidermal growth factor, is passively transferred from a dilute aqueous solution. When placed in contact with a wounded tissue, the growth factor passively transfers out of the contact lens to provide accelerated healing. The amount of growth factor absorbed into the hydrogel is low, e.g., ≦350 ppb, but this amount has surprisingly been found to be effective in producing a therapeutic effect. The systems are applicable to ocular and other wound treatments.

BACKGROUND OF THE INVENTION

[0001] In general, the invention relates to the fields of hydrogels,drug delivery systems, and wound healing.

[0002] Corneal wounds caused by injury, disease, or surgery represent aserious medical condition that may lead to loss of sight. For example,persistent epithelial defects can lead to stromal melting, which causesserious visual dysfunction. Wound healing of corneal mucosal tissue hastaken on increased importance with the advent of laser correctivesurgery to re-establish normal vision for people who do not wish to wearcontact lenses or spectacles. These laser surgical methods are used tocorrect vision for nearsightedness (myopia), farsightedness (hyperopia),and astigmatism. The methods include laser in situ keratomileusis(LASIK), laser epithelial keratomileusis (LASEK), and photo-refractivekeratectomy (PRK).

[0003] LASIK refers to the use of a laser to reshape the cornea withoutinvading the adjacent cell layers. During the LASIK procedure, amicrokeratome is used to separate the surface layers of the cornea andcreate a corneal flap (160-180 microns deep). This flap stays attachedto the rest of the cornea and is folded back on one side to expose thestroma of the cornea. The laser delivers pulses of ultraviolet lightonto the inner cornea (stroma). Each pulse removes a microscopic layerof the inner cornea to reshape the surface of the cornea. Fornearsighted patients, the procedure flattens the cornea. For farsightedpatients, the procedure increases the curvature of the cornea. Forastigmatism, selected tissues are removed at certain angles to make thecornea more spherical in shape. After exposure to the laser iscompleted, the corneal flap is replaced where it bonds without the needfor stitches. The anterior layers of the cornea (epithelium, Bowman'sLayer) are largely preserved. Once the surgery is completed, the eye isleft to heal normally with the exception of eye drops, which are used toprevent infection & swelling, with varying degrees of success. Followingthe surgery, patients are able to see clearly without depending onglasses or contacts.

[0004] During PRK, the surgeon removes the epithelium (the anteriorlayer of the cornea or Bowman's Layer), which is a thin layer ofprotective skin that covers the cornea. This layer can be removed withan excimer laser or a brush. During the procedure, the patient stares ata fixation light. In less than a minute, the laser removes the properamount of tissue while it reshapes the surface of the cornea. Theexcimer laser delivers pulses of ultraviolet light into the cornea. Thisexposure to laser radiation reduces or eliminates nearsightedness byflattening the central cornea and relocating the focal point of the lensonto the retina rather than in front of it, which produces sharpervision. Following surgery, a bandage contact lens is placed on the eyefor 2-3 days. Because the epithelium was removed, patients mayexperience blurry vision for three to five days. Eye drops and thecontact lens are effective in reducing postoperative discomfort. Thepurpose of the contact lens given to PRK patients post-surgically is toprotect the leading edge of the corneal epithelium that is regeneratingalong the surface of the eye, post-surgery. As patients blink, the newerleading edge of the epithelium may be removed. As a result, recoverytakes longer and there is an increased risk of infection.

[0005] LASEK is similar to PRK but the epithelium is detached by usingan alcohol solution that weakens the epithelium and allows it to foldback into a flap. A laser is then used to re-shape the cornea andcorrect vision acuity.

[0006] All three procedures can result in corneal epithelial defects,and inflammation and infection may also occur. These complications canlead to acuity regression or other adverse effects. Wound healing isthus of critical importance for the outcome of surgery. There exists aneed, therefore, for devices that promote healing of corneal wounds.

SUMMARY OF THE INVENTION

[0007] The present invention features hydrogel drug delivery systems andmethods of producing and using such systems for the treatment of wounds.The systems are based on a hydrogel into which a growth factor, e.g.,epidermal growth factor (EGF), is passively transferred from a diluteaqueous solution. When placed in contact with a wounded tissue, thegrowth factor passively transfers out of the hydrogel to provideaccelerated healing and a concomitant reduction in pain. The amount ofgrowth factor absorbed into the hydrogel is low, e.g., ≦350 ppb, butthis amount surprisingly is effective in producing a therapeutic effectlikely because the delivery system is localized and provides a sustainedrelease of the factor. The systems are applicable to ocular wounds,especially after vision correcting surgery, as well as other woundtreatments.

[0008] Accordingly, in one aspect, the invention features a polymerichydrogel that comprises a substantially pure growth factor, e.g.,epidermal growth factor, platelet derived growth factor, hepatocyticgrowth factor, or combinations thereof, at a concentration of between 5ppb and 350 ppb. The hydrogel has a water content of, for example,between 37.5% and 70% by weight. Exemplary hydrogel materials include atetrapolymer of hydroxymethylmethacrylate, ethylene glycol,dimethylmethacrylate, and methacrylic acid. Other examples of hydrogelsinclude those including an ionic or non-ionic polymer and thoseincluding etafilcon A, vifilcon A, or polymacon B. In variousembodiments, the growth factor is capable of being passively releasedinto an environment, e.g., an ocular environment, under ambient orexisting conditions. In other embodiments, the hydrogel may be shaped asa contact lens, e.g., one capable of correcting vision. Such a contactlens may be capable of correcting vision in the range of +8.0 to −8.0diopters, including plano, and may have a base curve between 8.0 and9.0.

[0009] The invention further features a method for making a hydrogeldrug delivery system. This method includes the steps of providing ahydrogel; washing the hydrogel in an isotonic saline solution; partiallydesiccating the lens; and placing the washed and partially desiccatedhydrogel in an aqueous solution, e.g., having a pH between 6.9 and 7.4,of between 0.01 and 10 ng growth factor, e.g., epidermal growth factor,platelet derived growth factor, hepatocytic growth factor, orcombinations thereof, per μl, wherein growth factor is passivelytransferred into the hydrogel to produce the hydrogel drug deliverysystem. The concentration of growth factor in the hydrogel after soakingis, for example, between 5 and 350 ppb. In one embodiment, the hydrogelis placed in the solution of growth factor for at least 30 minutes.

[0010] In another aspect, the invention features a method for treating awound. The method includes the steps of providing a hydrogel containinga growth factor as described above; placing said hydrogel in contactwith the wound, wherein the growth factor is passively released from thehydrogel to treat the wound. In one embodiment, the hydrogel furtheracts as a protective shield against mechanical abuse. In variousembodiments, the wound is in endothelial tissue, epithelial tissue, thelung, the skin, or the digestive tract. The hydrogel may be placed in abody cavity. In another embodiment, the passively released growth factorcauses a reduction in pain compared to a wound not contacted with thepolymeric hydrogel.

[0011] By “ambient conditions” is meant room temperature and pressure.

[0012] By “existing conditions” is meant in situ, as in the eye or otherbody system.

[0013] By “substantially pure” is meant having a purity of greater than75% by weight. A growth factor of the invention is, for example, greaterthan 85%, 90%, 95%, or even 99% pure. Use of the term is intended todefine purity from other biological compounds, e.g., proteins,carbohydrates, and lipids that are commonly associated with the growthfactor in vivo.

[0014] By “treating” is meant the medical management of a patient withthe intent that a prevention, cure, stabilization, or amelioration ofthe symptoms will result. This term includes active treatment, that is,treatment directed specifically toward improvement of the disorder;palliative treatment, that is, treatment designed for the relief ofsymptoms rather than the curing of the disorder; preventive treatment,that is, treatment directed to prevention of the disorder; andsupportive treatment, that is, treatment employed to supplement anotherspecific therapy directed toward the improvement of the disorder. Theterm “treatment” also includes symptomatic treatment, that is, treatmentdirected toward constitutional symptoms of the disorder. The termfurther includes the promotion of wound closure or healing.

[0015] By “wound” is meant an injury to any tissue. Examples of woundsinclude burns, lacerations, abrasions, bites, surgical wounds, puncturewounds, and ulcers.

[0016] All percentages described in the present invention are by weightunless otherwise specified.

DETAILED DESCRIPTION OF THE INVENTION

[0017] This invention provides a polymeric drug delivery systemincluding a hydrogel containing a growth factor, e.g., EGF. Allowingpassive transference of the growth factor from a dilute aqueous solutioninto the hydrogel produces the delivery system. The hydrogel, whenplaced in contact with a wound, delivers a low concentration of thegrowth factor over an extended period of time. This sustained deliveryaccelerates the wound healing process while avoiding potential damagingeffects of localized delivery of high concentrations of compounds.

[0018] Drug Delivery System

[0019] Hydrogels. This invention may employ different polymercompositions that are useful in the treatment of a variety of tissues.For example, in the ocular environment, conventional soft contact lensescan be used and can be either ionic or non-ionic hydrogels containingbetween 37.5% -70% water by weight and can have any base curve, e.g.from 8.0 to 9.0. The contact lenses may also have the ability to correctvision, for example over a range of diopters of +8.0 to −8.0, includingpiano. Exemplary hydrogel contact lens materials include etafilcon A,vifilcon A, polymacon B, and a tetrapolymer ofhydroxymethylmethacrylate, ethylene glycol, dimethylmethacrylate, andmethacrylic acid. These materials may also be employed, in otherphysical forms, in treating wounds in other tissues. Other suitablehydrogel materials are known to those skilled in the art. The hydrogelsmay be insoluble or may dissolve over time in vivo, e.g., over one dayor one week. The growth factor is passively delivered, for example, bydiffusion out of the hydrogel or by release as the hydrogel dissolves.

[0020] The drug delivery system is produced by partially desiccating ahydrated hydrogel (or equivalently partially hydrating a desiccatedhydrogel). The desiccation step removes, for example, approximately(within ±2%) 5%, 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60%, or 75% of thewater in a hydrogel. Desiccation can occur, for example, by exposure ofthe hydrogel to ambient or humidity controlled air, by heating thehydrogel for a specific period of time, or by blowing dried gas, such asN₂, over the hydrogel. In one embodiment, the hydrogel is saturated withphysiological (isotonic) saline prior to desiccation. The partiallydesiccated hydrogel is then soaked, e.g., for at least 30 minutes, in adilute aqueous solution of growth factor, e.g., at a pH between 6.9 to7.4. The hydrogels may also be soaked for at least 1 hour, 6 hours, 12hours, or 24 hours. The concentration of growth factor into which thehydrogel is placed is typically 10 ng/μl or less, e.g., at most 5 ng/μl,1 ng/μl, 0.1 ng/μl, or 0.01 ng/μl. Higher concentrations may also beused, for example, to reduce the soaking time. The growth factor ispassively transfered into the hydrogel. This transfer occurs at least inpart by rehydrating the hydrogel. Diffusion of the growth factor intothe water in the hydrogel may also occur.

[0021] Typically, the concentration of growth factor transferred to thehydrogel is substantially lower than the solution in which the hydrogelis soaked. For example, the concentration of growth factor in thehydrogel is at least 2×, 5×, or 10× less than that of the soakingsolution. The water content and type of hydrogel, time and conditions,e.g., temperature of soaking, composition of the soaking solution (e.g.,ionic strength and pH), and type of growth factor employed also mayinfluence the concentration of growth factor in the drug deliverysystem. The water content of the hydrogel also helps to determine thetotal amount of growth factor present in a hydrogel. Thus, the watercontent of a hydrogel represents another variable by which to controlthe amount of growth factor delivered to a tissue. The production of ahydrogel containing a specified amount of growth factor can beaccomplished by routine experimentation by one skilled in the art.Exemplary hydrogels include between 5 and 350 ppb of growth factor, forexample, between 5 and 250 ppb, 5 and 100 ppb, 5 and 50 ppb, or 5 and 10ppb.

[0022] Growth factors. Growth factors are a heterogeneous group ofproteins capable of stimulating growth and the multiplication of cells.Exemplary growth factors include epidermal growth factor, plateletderived growth factor, hepatocytic growth factor, and combinationsthereof. These growth factors may be natural, synthetic, or recombinantgrowth factors or growth factor derivatives from any animal, forexample, humans, or any domesticated animal or pet species. Such growthfactors also include biologically active growth factors and analogs.Peptide growth factors play important biological roles by regulatingmany of the processes involved in normal wound healing includingmigration, mitosis, and differentiation of cells. Growth factors arecommercially available or may be isolated using methods known in theart.

[0023] Other compounds. The hydrogels of the invention may also containmedicaments other than growth factors. These additional compoundsinclude, without limitation, analgesics, anti-inflammatory drugs,antibodies, meganins, self-proteins, pharmaceutical drugs, andantibiotic compounds.

[0024] The use of preservatives is non-ideal as they may transfer to ahydrogel disproportionately and cause cytotoxicity.

[0025] Treatment. To treat a wound, a drug delivery system of theinvention may be placed in contact with a damaged tissue. When thesystem is shaped as a contact lens, the lens may simply be placed in theeye normally in order to deliver the growth factor. In order to effectaccelerated healing of other wounds, the hydrogel may be part of abandage or may be adhered (e.g., by adhesives or sutures) to the woundedtissue. If the hydrogel is placed internally in a patient, the hydrogelis advantageously biodegradable.

[0026] Hydrogels may be considered to be disposable and may be replacedafter a specified period of time. Alternatively, a hydrogel that has adepleted amount of growth factor may be recycled by desiccating andsoaking the hydrogel again.

[0027] Treatment Approaches

[0028] The invention may be used in conjunction with healing many typesof wounds, including, without limitation, ocular, oral, lung, digestivetract, skin, large intestine, small intestine, colon, and other woundsto endothelial, mucosal, or epithelial tissues. As stated above, theinvention provides accelerated healing by delivering a growth factor toan injured tissue. This delivery occurs by passive transfer and allowsmedications to be released into fluids of the body, e.g., ocular fluid.The growth factor stimulates proliferation of cells surrounding a woundto close the wound and replace damaged cells. Because the growth factoris localized by the hydrogel, which provides a timed release of thegrowth factor, a lesser amount of growth factor may in many cases beneeded to effect wound healing than if, e.g., topical solutions, such aseye drops are used. Accelerated healing may also reduce the pain andinflammation associated with a particular wound and may help preventinfection. In addition, the hydrogel may also act as a physical barrierto provide protection from mechanical abuse and to prevent adherence ofthe healing tissue to adjacent tissues.

[0029] Ocular Wounds. In one embodiment, the wound is an ocular wound,e.g., in epithelial, endothelial, or retinal tissue. The invention is ofparticular utility after vision correcting surgery, such as LASIK, PRK,or LASEK. Soft and collagen contact lenses may be utilized to minimizepost-surgical epithelial trauma and provide a stable healingenvironment. PRK typically requires a therapeutic contact lens for 3-4days, and post-operative therapeutic drops are often prescribed. In thepresent invention, the hydrogel may be shaped as a contact lens thatacts as a reservoir for the growth factor and can serve to protect theleading edge of wound healing from normal mechanical abuse. The growthfactor gradually delivered in a low concentration from the lens obviatesthe need for therapeutic drops. Therapeutic drops often include highconcentrations of drugs because the majority of the drop is excretedfrom the eye in a short period of time. These high concentrations cancause additional damage to a wound, which is avoided by the use of thepresent, localized time-release drug delivery system.

[0030] A further understanding of the invention may be obtained from thefollowing non-limiting examples.

EXAMPLE 1

[0031] Production of a Drug Delivery System.

[0032] An exemplary drug delivery system was prepared as follows.Contact lenses were removed from their package and rinsed with saline toremove contact lens packing solution. The hydrogel lens materials wereallowed to desiccate for 10-30 seconds. The hydrogel lens materials wereplaced into physiological saline that contained epidermal growth factor(EGF) at concentrations of 10 ng/μl or 5.0 ng/μl for at least 30minutes. Lower concentrations may also be used. Longer passivetransference times may also be used. Untreated or control lenses wereplaced in physiological saline without EGF.

EXAMPLE 2

[0033] Healing of Ocular Tissue.

[0034] Ocular cells were placed into a sterile plastic dish. This dishcontained a 5-mm disk. The purpose of the disk was to prevent cells fromgrowing in the covered area. When the disk was removed, a 5-mm “wound”or “hole” was present.

[0035] Contact lenses were then added to these cell sheets with thewounds. The lenses were left in contact with the cell sheets for aminimum of 30 minutes. Minimal medium was used to maintain the cellcultures. Cells were incubated at 35° C.±2° C. in 5% CO₂. Contact lenseswith or without EGF were produced as in Example 1. The contact lensesused were polymacon B, vifilcon A, and lidofilcon A hydrogel polymers.

[0036] The cell sheets were then viewed over time, and the diameter ofthe hole was measured.

[0037] The results are expressed in terms of closure of the in vitrowound over time.

[0038] Epithelial Cells and Tissue. Epithelial (rabbit cornealepithelial cells) cells were seeded on a dish and contacted with controland EGF-containing contact lenses. At 48 hours there was a 25%difference in the closure rate between the EGF-treated cells and thenon-EGF treated cells. At 72 hours, there was a 43% difference in theclosure rate between the EGF-treated epithelial tissue and the controls.The hydrogel material that was used was vifilcon A, an ionic polymerwith a water content of 55%. The polymer had been incubated with 10ng/μl for one hour at 4° C. prior to use in the experiments. The lenscontained approximately 1 ng/μl of EGF.

[0039] Closure rates were calculated by direct measurement of thediameter of the wound. Measurements were taken daily.

[0040] In a related series of experiments, a vifilcon A lens wasincubated under the same conditions as above with 5.0 ng/μl of EGF andthen contacted with an epithelial “wound” as above. The lens containedapproximately 0.5 ng/μl of EGF. At 48 hours, there was a 21% closurerate difference between controls and EGF treated hydrogel materials. At72 hours, there was also a 21% difference in the closure rate. Theseresults indicated that over a 72-hour period, the relative healing ratesremained essentially the same for the treated and non-treated epithelialtissue, with the epithelial tissue treated with EGF always having anaccelerated rate of healing.

[0041] The rate of wound healing increased with increased exposure ofthe hydrogel material to the wound. Further, compared to a wound notcontacted with any lens, at 48 hours there was a 31% difference in thehealing rates. Healing for tissue exposed to a lens soaked in 10 ng/μlof EGF increased from 14% at 48 hours to 25% at 72 hours.

[0042] Endothelial Cells and Tissue. Wounds caused in endothelial tissue(bovine corneal endothelial cells) were also healed by release of EGFfrom a vifilcon A lens. The lens, soaked in 10 ng/μl of EGF as above,showed a 73% difference in healing rates at 48 hours compared to acontrol. At 72 hours, the EGF-treated tissue had completely healed. Inthe control group, less than half (43%) of the tissue had healed. Thesame lens material exposed to 5 ng/μl of EGF showed a 31% difference inclosure rate at 48 hours between the EGF treated group and the controls.At 72 hours, 53% of the tissue had healed in the EGF treated group,compared to 43% in the control.

[0043] Lidofilcon A hydrogel (non-ionic, water content=70%) materialswere evaluated for their ability to deliver EGF to endothelial tissue toclose wounds. The concentration of EGF used in the soaking solution was10 ng/μl. At 48 hours, the EGF treated tissue showed a 54% increase inthe healing rate (wound closure rate) as compared to controls. At 72hours, there was a difference of 44%.

[0044] A third material, Polymacon B, that is non-ionic and has a watercontent of 38%, was also evaluated for the ability to deliver EGF towounds. The lenses were prepared using a soaking solution of 10 ng/μl ofEGF. At 48 hours, the wound was 60% closed in the treated group and 27%closed in the non-treated group. At 72 hours, the difference in closurebetween the treated and untreated groups was 62%. In the EGF treatedgroup at 72 hours, the wound had closed by 80%, while in the untreatedgroup, the wound had closed by 46.8%.

[0045] Other Embodiments

[0046] Modifications and variations of the described methods of theinvention will be apparent to those skilled in the art without departingfrom the scope and spirit of the invention. Although the invention hasbeen described in connection with specific desirable embodiments, itshould be understood that the invention as claimed should not be undulylimited to such specific embodiments. Indeed, various modifications ofthe described modes for carrying out the invention, which are obvious tothose skilled in the art, are intended to be within the scope of theinvention.

[0047] All publications, patents, and patent applications mentioned inthis specification are herein incorporated by reference to the sameextent as if each individual publication, patent, or patent applicationwas specifically and individually to be incorporated by reference.

[0048] Other embodiments are within the claims.

What is claimed is:
 1. A polymeric hydrogel that comprises asubstantially pure growth factor at a concentration of between 5 ppb and350 ppb.
 2. The hydrogel of claim 1, wherein said growth factor isselected from the group consisting of epidermal growth factor, plateletderived growth factor, hepatocytic growth factor, and combinationsthereof.
 3. The hydrogel of claim 2, wherein said growth factor isepidermal growth factor.
 4. The hydrogel of claim 1, said hydrogelcomprising a water content between 37.5% and 70% by weight.
 5. Thehydrogel of claim 1, said hydrogel comprising a tetrapolymer ofhydroxymethylmethacrylate, ethylene glycol, dimethylmethacrylate, andmethacrylic acid.
 6. The hydrogel of claim 1, wherein said growth factoris capable of being passively released into an environment under ambientconditions.
 7. The hydrogel of claim 6, wherein said environment is anocular environment.
 8. The hydrogel of claim 1, wherein said growthfactor is capable of being passively released into an environment underexisting conditions.
 9. The hydrogel of claim 8, wherein saidenvironment is an ocular environment.
 10. The hydrogel of claim 1,wherein said hydrogel is shaped as a contact lens.
 11. The hydrogel ofclaim 10, wherein said hydrogel is capable of correcting vision.
 12. Thehydrogel of claim 11, wherein said hydrogel is capable of correctingvision in the range of +8.0 to −8.0 diopters, including plano.
 13. Thehydrogel of claim 10, said hydrogel having a base curve between 8.0 and9.0.
 14. The hydrogel of claim 1, said hydrogel comprising an ionicpolymer.
 15. The hydrogel of claim 1, said hydrogel comprising anon-ionic polymer.
 16. The hydrogel of claim 1, said hydrogel comprisingetafilcon A, vifilcon A, or polymacon B.
 17. A method for making ahydrogel drug delivery system, said method comprising the steps of: (a)providing a hydrogel; (b) washing the hydrogel in an isotonic salinesolution; (c) partially desiccating the lens; and (d) placing the washedand partially desiccated hydrogel in an aqueous solution of between 0.01and 10 ng growth factor per μl, wherein growth factor is passivelytransferred into said hydrogel to produce said hydrogel drug deliverysystem.
 18. The method of claim 17, wherein the concentration of growthfactor in said hydrogel after step (d) is between 5 and 350 ppb.
 19. Themethod of claim 17, wherein said growth factor is selected from thegroup consisting of epidermal growth factor, platelet derived growthfactor, hepatocytic growth factor, and combinations thereof.
 20. Themethod of claim 19, wherein said growth factor is epidermal growthfactor.
 21. The method of claim 17, wherein said aqueous solution instep (d) has a pH between 6.9 and 7.4
 22. The method of claim 17,wherein said hydrogel is shaped as a contact lens.
 23. The method ofclaim 17, wherein, in step (d), said hydrogel is placed in said solutionfor at least 30 minutes.
 24. A method for treating a wound, said methodcomprising the steps of: (a) providing a polymeric hydrogel comprisingbetween 5 and 350 ppb by weight of growth factor; and (b) placing saidhydrogel in contact with said wound, wherein said growth factor ispassively released from said hydrogel to treat said wound.
 25. Themethod of claim 24, wherein said growth factor is selected from thegroup consisting of epidermal growth factor, platelet derived growthfactor, hepatocytic growth factor, and combinations thereof.
 26. Themethod of claim 25, wherein said growth factor is epidermal growthfactor.
 27. The method of claim 24, wherein said wound is in an eye andsaid hydrogel is shaped as a contact lens.
 28. The method of claim 24,wherein said hydrogel further acts as a protective shield againstmechanical abuse.
 29. The method of claim 24, wherein said wound is inendothelial tissue.
 30. The method of claim 24, wherein said wound is inepithelial tissue.
 31. The method of claim 24, wherein said wound is alung, skin, or digestive tract wound.
 32. The method of claim 24,wherein, in step (b), the hydrogel is placed in a body cavity.
 33. Themethod of claim 24, wherein said passively released growth factor causesa reduction in pain compared to a wound not contacted with saidpolymeric hydrogel.